In article <4ihofq$kvp at ccshst05.cs.uoguelph.ca> wyu at uoguelph.ca (Wenjin Yu) writes:
>From: wyu at uoguelph.ca (Wenjin Yu)
>Subject: SOS!--PCR restriction cut
>Date: 17 Mar 1996 19:17:14 GMT
> I have a big problem to cut PCR product in which the restriction site
>is generated in the primer. At first I designed a primer containing a Bam
>HI site. Based on the inforamtion in New England Biolabs Catalog, 4
>extra bases was added at the 5'-end, but the PCR product couldn't be cut
>no matter how much enzyme was used. Then I made another primer
>containing an EcoRI site, and 15 extra-bases was added to the 5'-end this
>time. But it seems still not work! I was wondering if anyone can help me to
>deal with this big trouble. Many thanks.
> (please send your HELP to my mail box: wyu at uoguelph.ca)
>Wenjin Yu
>U of Guelph
I'd recommend you try another enzyme supplier. I have had problems digesting
PCR products with NEB enzymes. I switched to Boehringher Mannheim and that
took care of my problems. This only happened with PCR products.
Hector "Che" Cruz-Lopez
Florida Game and Fish Commission
Boca Raton, Florida